1. Field of the Invention
This invention relates to methods of preparing 7-substituted camptothecin compounds and 7-substituted camptothecin analogs. Camptothecin (CPT) and CPT analogs have been reported to inhibit the enzyme topoisomerase I and have in vitro and in vivo anticancer activity. It has been shown that a large number of substituents can be placed at C7 of CPT without loss of activity (Redinbo et al., Science 279, 1504-1513, 1998).
2. Background of the Invention
Camptothecin (CPT) is a naturally occurring cytotoxic alkaloid which is known to inhibit the enzyme topoisomerase I and is a potent anti-tumor agent. Camptothecin compounds have the general ring structure shown below. Camptothecin was isolated from the wood and bark of Camptotheca acuminata by Wall et al. (Wall et al., 1966, J. Am. Chem. Soc., 88:3888).
Major synthetic efforts have been directed to derivatizing the B-ring at C7 to improve cytotoxic and in vivo activity.
The cytotoxic activity of camptothecin compounds is believed to arise from the ability of these compounds to inhibit both DNA and RNA synthesis and to cause reversible fragmentation of DNA in mammalian cells. Topoisomerase I relaxes both positively and negatively supercoiled DNA and has been implicated in various DNA transactions such as replication, transcription and recombination. The enzyme mechanism is believed to involve a transient breakage of one of the two DNA strands and the formation of a reversible covalent topoisomerase I enzyme-DNA complex. Camptothecin interferes with the DNA breakage-reunion reaction by reversibly trapping the enzyme-DNA intermediate termed the “cleavable complex.” The cleavable complex assay is a standard test for determining the potential and in vivo cytotoxic activity of camptothecin compounds. The high levels of topoisomerase I in several types of human cancer and the low levels in correspondingly normal tissue provide the basis for tumor treatment with biologically active camptothecin analogs.
U.S. Pat. No. 4,894,456 describes methods of synthesizing camptothecin compounds which act as inhibitors of topoisomerase I and are effective in the treatment of leukemia (L-1210). U.S. Pat. No. 5,225,404 discloses methods of treating colon tumors with camptothecin compounds.
Numerous camptothecin compounds and their use as inhibitors of topoisomerase I are reported by U.S. Pat. Nos. 5,053,512; 4,981,968; 5,049,668; 5,106,742; 5,180,722; 5,244,903; 5,227,380; 5,122,606; 5,122,526; and 5,340,817.
U.S. Pat. No. 4,943,579 discloses the esterification of the hydroxyl group at the 20-position of camptothecin to form several prodrugs. This patent further discloses that the prodrugs are water soluble and are converted into the parent camptothecin compounds by hydrolysis.
Wall et al. U.S. Pat. Nos. 5,646,159 and 5,916,892 disclose C20 amino acid esters of CPT compounds.
Wall et al. U.S. Pat. No. 5,932,588 disclose CPT compounds bearing a C7 methylene leaving groups at C7 such as —CH2L where L is Cl, Br, I, —OSO2CH3, —OSO2C6H4—CH3, etc.
Brangi et al., Cancer Research, 59, 5938-5946 Dec. 1, 1999, report an investigation of Camptothecin resistance in cancer cells and report the compound difluoro-10,11-methylenedioxy-20(S)-camptothecin and several C7-substituted compounds.
A need continues to exist, however, for a method of preparing 7-substituted camptothecin compounds. Refs: Du et al., Biorg. and Med. Chem. 10, 103-110 (2002); Dallavalle et al., J. Med. Chem. 44, 3264-3274 (2001).
The procedure of Sawada et al., Chem. Pharm. Bull. 39, 2574-2580 (1991) for preparing 7-alkyl compounds gives adequate yields for C1-3 alkyl compounds; however, the yields for C4, C5, C6-alkyl rapidly become poor. We have discovered a novel way of preparing a large variety of alkyl and aryl C7-substituted compounds in excellent yields by the reaction of an orthoaminobenzonitrile or appropriately substituted orthoaminobenzonitrile with a large variety of organometallic reagent which will be described in detail in this application.